采用激光共聚焦扫描显微镜技术、免疫印迹技术和流式细胞术3种方法对鼠抗三疣梭子蟹血细胞多克隆抗体与黄道蟹、日本蟳、中华绒螯蟹、勘察加帝王蟹血细胞的免疫交叉反应进行分析.激光共聚焦扫描显微镜结果表明,该抗体与黄道蟹、日本蟳、勘察加帝王蟹、中华绒螯蟹血细胞均发生了免疫交叉反应,且发生反应的抗原决定簇均位于细胞膜上;流式细胞术测定该抗体与黄道蟹颗粒血细胞和透明血细胞交叉反应阳性率最高,分别为99.45%、98%,与中华绒螯蟹颗粒血细胞和透明血细胞交叉反应阳性率最低,阳性率分别为81.34%、78.56%;免疫印迹结果显示该抗体与日本蟳血细胞结合的蛋白分子量是80、38.5 ku;与勘察加帝王蟹血细胞结合的蛋白分子量是80、93 ku;与中华绒螯蟹血细胞结合的蛋白分子量是82、41、30 ku;与黄道蟹血细胞结合的蛋白分子量是70、43 ku.%In order to research the role of hemocytes in immune mechanism of crustaceans, polyclonal antibodies against hemocytes of swimming crab Portunus trituberculatus were raised, and laser scanning confocal microscopy (LSCM), Western-blotting and Flow Cytometry (FCM) were used to detect the cross-reaction between antisera of swimming crab and 4 species crustacean (Cancer magister,Charybdis japonic, Eriocheir sinensis, Paralithodes camtschaticus)hemocytes.Two types of hemocytes in C.japonic,E.sinensi, and P.camtschaticus and three types of hemocytes in C.magister were distincted by FCM.The strongest intensity of immuno-reactivity was in C.magister by LSCM, and the maximal positive rate was that antisera reacted with hemocytes of C.magister detected by FCM, with 99.45% of granulocytes and 98% of hyalinocytes.The results indicated antigenic similarities were found among the two species.Thus, C.magister being more closely related species to swimming crab exhibited many immuno-reactions, while the other species with a phylogenetically longer distance to swimming crab did not.The immuno-reactive epitopes in C.magister and swimming crab on the hemocyte membranes and in the plasma seemed to be well conserved, however, they are not necessarily the same, with similar functions.There was higher positive rate in granulocytes (81.34%) than in hyalinocytes (78.56%) detected by FCM in four species.The findings indicate that antigenic similarities exist among the four crustaceans species, C.magister being more closely related species to swimming crab.
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